The locus coerulus (LC) is a small brainstem nucleus that is the primary source of norepinephrine (NE) to the cortex, thalamus, midbrain, cerebellum, and spinal cord. The LC-NE system is thought to play important roles in many normal brain functions, including arousal and sensory-motor processing. In addition, the LC-NE system has been implicated in a number of clinical disorders, including attention-deficit/hyperactivity disorder (ADHD), anxiety, depression, and schizophrenia. Thus, a better understanding of the properties of the LC-NE system is likely to provide important insights into both normal and abnormal brain function. A major obstacle to such research is the size and location of the LC, which make it difficult to target for recording or imaging. One possible solution to this problem is to measure pupil diameter, which is relatively straightforward to measure and under certain low-light conditions is thought to reflect LC activation. However, despite some strong claims to the contrary, the relationship between LC activation and pupil diameter has never been established with sufficient rigor to allow pupillometry measurements to be interpreted in terms of LC function. The goal of the proposed studies is to characterize this relationship in detail. We will use three complementary approaches: 1) simultaneous recording of individual LC neurons and pupil diameter in awake, behaving monkeys under a variety of conditions;2) simultaneous application of electrical microstimulation in the LC and measurements of pupil diameter in awake, behaving monkeys;and 3) measurements of pupil diameter in human subjects under comparable conditions to those tested with monkeys. Together, these studies will provide by far the most rigorous assessment to date of the relationship between LC activation and pupil diameter. Given the growing interest in the role of the LC-NE system in behavior, cognition, and disease, and the relative ease with which pupil diameter can be measured, these results are highly likely to make a strong impact on the design and interpretation of many future studies. PUBLIC HEALTH RELEVANCE: The proposed work is designed to test, rigorously and for the first time, a proposed link between pupil diameter and the activation of brainstem neurons in the locus coeruleus (LC) that are the primary source of norepinephrine (NE) to the rest of the brain. This work will establish a solid foundation for future uses of non-invasive pupil measurements to better understand normal LC function and to help diagnose and treat the many disorders associated with abnormal LC function, including ADHD, anxiety, depression, and schizophrenia.